Phenylketonuria (PKU) is an inherited metabolic disease of humans caused by the deficiency of, or low activity of, the enzyme phenylalanine hydroxylase. Phenylalanine hydroxylase normally functions in the human body to convert the amino acid phenylalanine to tyrosine. When this enzyme is deficient, phenylalanine and its abnormal breakdown products accumulate in the bloodstream. These breakdown products are harmful to developing cells of central nervous system, resulting in symptoms of mental retardation, developmental delay, and seizures. PKU is caused by recessive mutations in a single allele in the homozygous state. About 1 in every 15,000 infants born in the United States is homozygous for this allele.
Dietary therapies have been used to effectively treat infants with PKU and prevent PKU symptoms from appearing. All states require routine tests of all newborn babies to detect PKU homozygotes. Those identified at birth are put on a special diet containing low amounts of phenylalanine. These diets provide enough phenylalanine (an essential amino acid) to supply dietary needs but not enough to permit toxic accumulations.
On the basis of 32 years of experience, it is now clear that administration of low phenylalanine diets, especially during at least the first fifteen years of life (when brain development is still in progress), allow PKU individuals to develop normally. However, PKU individuals must continue to maintain a diet low in phenylalanine throughout their entire life to avoid intellectual impairment and behavioral or psychological disorders caused by elevated blood levels of phenylalanine.
Dietary compliance of a pregnant PKU female is particularly important to ensure normal development of her child. Since phenylalanine is selectively concentrated by the placenta in the amniotic fluid of a pregnant female, maternal blood levels as low as 3-4 times normal levels can harm the developing fetus and lead to birth defects (heart and brain defects). The occurrence of birth defects arising from the mother's elevated phenylalanine levels is typically referred to as Maternal PKU Syndrome. To prevent Maternal PKU Syndrome, a PKU mother-to-be must be placed on a low phenylalanine diet prior to conception.
Dietary therapy using phenylalanine-deficient amino acid mixtures was first reported in 1953 by Bickel et al. Current treatment focuses upon the use of a low-phenylalanine diet consisting of a special metabolic formula with supplemental low-protein foods. The metabolic formulas are composed of crystalline amino acids, fats, carbohydrate sources, trace minerals and vitamins to provide a food with high amino acid content that is low or totally deficient in phenylalanine. The total dietary phenylalanine intake of a phenylketonuria individual receiving the special formulas is determined by the content of the table foods other than formula. Since dietary treatment is mandatory for the life of the individual and all high-quality protein has a substantial content of phenylalanine, intake of meats and dairy products is prohibited. Unfortunately, the administration of metabolic formulas to PKU individuals is complicated by poor palatability and odor, limited solubility of certain amino acids, and inadequate resistance to thermal degradation associated with baking or cooking.
Previous workers have altered the amino acid composition of the formulas to reduce the content of methionine and increase the amount of glutamine to improve the taste and odor, in an attempt to enhance not only the palatability of the product but also the social acceptance of medical food by family members. Other producers have also packaged the formula in fruit bar form to help disguise the taste.
The present invention solves the problem of poor palatability, taste, and processibility by utilizing an intact phenylalanine-deficient protein that supplies most of the required essential amino acids in a form (a protein) that does not have an appreciably bad taste or odor. The phenylalanine-deficient proteins of the present invention are intact natural proteins, most typically native plant proteins, that have been modified, other than by hydrolysis, to reduce or eliminate phenylalanine from the amino acid sequence of the protein. The term "intact protein" as used in accordance with the present invention is defined as an unhydrolyzed natural/native protein comprising a continuous series of amino acids linked together through peptide bonds. The term is used to distinguish the present phenylalanine deficient protein compositions over compositions comprising hydrolyzed or partially hydrolyzed proteins. The intact, modified natural proteins of the present invention may be used as a supplement in formulas or combined with protein deficient products and cooked in a manner similar to traditional foods.
Advantageously, the intact modified proteins retain many physical properties of the parent protein, including solubility characteristics, low odor and minimal taste, which allow it to be mixed into liquids, baked, or cooked in traditional mixtures. These properties make the protein products of the present invention superior to crystalline amino acid formulations, fractionated protein hydrolysis products or synthetic polypeptides.
One object of the present invention is to modify the amino acid sequence of a natural protein to reduce or eliminate phenylalanine from the amino acid sequence of the protein. These intact, modified natural proteins provide a protein supplement for use in dietary therapies for PKU patients to minimize the physiological symptoms of PKU.
An additional object of the present invention is to provide a method for synthesizing phenylalanine free protein.
Many protein sources are nutritionally incomplete in that they lack one or more of the essentials amino acids for proper nutrition of higher animals. A further object of the present invention is to increase the nutritional content of natural proteins by modifying the amino acid content of the intact protein to provide a protein that contains all the essential amino acids except phenylalanine.